Polyacetylenes from the Roots of Swietenia macrophylla King

Abstract

A phytochemical investigation of the roots of Swietenia macrophylla led to the isolation of seven polyacetylenes, including five new compounds (1–5) and two known ones (6–7). Their structures were elucidated by extensive spectroscopic analysis and detailed comparison with reported data. All the isolates were tested for their cytotoxicity against the human hepatocellular carcinoma cell line BEL-7402, human myeloid leukemia cell line K562, and human gastric carcinoma cell line SGC-7901. Compounds 1 and 6 showed moderate cytotoxicity against the above three human cancer cell lines with IC₅₀ values ranging from 14.3 to 45.4 μM. Compound 4 displayed cytotoxicity against the K562 and SGC-7901 cancer cell lines with IC₅₀ values of 26.2 ± 0.4 and 21.9 ± 0.3 μM, respectively.

1. Introduction

Natural polyacetylenes, structures featuring two or more triple bonds [1], are mainly found in plants belonging to the Araliaceae [2,3,4,5,6,7], Asteraceae [8,9,10], Umbelliferae [11,12,13], Santalaceae [14], Pittosporaceae [15], and Oleaceae [16] families. However, polyacetylenes are uncommon in the Meliaceae family, with only six such compounds having been found in total in this family, one of which is from Swietenia mahagoni [17], three from Toona ciliate [18], and two from the stem bark of Khaya ivorensis A. Chev [19,20]. Naturally occurring polyynes are classified into four types: acyclic C₁₈–C₁₄ acetylene compounds; acyclic C₁₃–C₈ acetylene compounds; compounds with an allene substructure; and aromatic and heterocyclic acetylene compounds [1]. Diverse polyacetylene structures exhibit a series of bioactivities, including cytotoxicity [8,10,11,21] and antimicrobial [14,22,23], antiviral [24], and enzyme-inhibitory [6,25] activities.

Swietenia macrophylla, a perennial deciduous timber tree that reaches a height of up to 50 m [26], is native to Central and South America [27] and widely distributed in West India, Malaysia, and southern China [28,29]. Antecedent chemical investigations on S. macrophylla have focused mostly on the aboveground parts and their bioactive limonoids [30]. It is necessary to expand the scope of research on S. macrophylla and discover or develop additional biologically active constituents of this plant genus [31]. Our recent study on the roots of S. macrophylla led to the isolation of a series of xanthones, limonoids, and other chemical components [32,33]. As a continuation of our studies on the biologically active agents from this plant, five new and two known acyclic C₁₈–C₁₄ polyacetylenes have been further isolated here, and their cytotoxic activities against the human hepatocellular carcinoma cell line BEL-7402, human myeloid leukemia cell line K562, and human gastric carcinoma cell line SGC-7901 were investigated. In this paper, the isolation, structural elucidation, and cytotoxicity of these compounds are reported as follows.

2. Results and Discussion

The chemical examination of the ethyl acetate (EtOAc) extract from the roots of S. macrophylla resulted in the isolation and identification of five new polyacetylenes (Figure 1), respectively named heptadeca-9-ene-4,6-diyne-3,11-diol (1), (E)-heptadeca-8-ene-4,6-diyne-3,10,11-triol (2), 10-methoxyheptadeca-4,6-diyne-3,9-diol (3), tetradeca-1,3-diyne-6,7,8-triol (4) and 6,7,8,9-tetraacetoxytetradeca-1,3-diyne (5), together with two known compounds, which were identified as α-hexy-3-(6-hydroxy-2,4-ocadiynyl)oxiranemethanol (6) [17] and (3R,8E,10S)-heptadec-8-ene-4,6-diyne-3,10-diol (7) [18] by comparing their experimental spectroscopic data with the reported data in the literature. HRESIMS and NMR spectra for compounds 1–5 are shown in the Supplementary Materials.

Compound 1 was obtained as a yellow oil with a positive optical rotation ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ +10 (c 0.1, CH₃OH) and has a molecular formula of C₁₇H₂₆O₂, as evidenced by the HRESIMS peak at m/z 285.1830 [M + Na]⁺ (calcd 285.1831 for C₁₇H₂₆NaO₂), requiring five unsaturation degrees. The UV spectrum displayed typical absorption bands for a conjugated ene-yne-yne chromophore at λ_max 280 and 270 nm [11], and the IR spectrum showed the OH group (3435 cm⁻¹), triple-bond (2234 cm⁻¹), and olefinic double-bond (1639 cm⁻¹) absorptions. The ¹H-NMR data (

Table 1. ¹H (500 MHz) and ¹³C-NMR (125 MHz) data for compounds 1–3 in CDCl₃ (δ in ppm, J in Hz).

Position	1		2		3
	δH	δC	δH	δC	δH	δC
1	0.99, t (7.4)	9.5, CH3	1.01, t (7.4)	9.5, CH3	1.01, t (7.4)	9.5, CH3
2	1.72, m	30.8, CH2	1.75, m	30.8, CH2	1.73, m	30.8, CH2
3	4.34, t (6.4)	64.1, CH	4.41, t (6.5)	64.3, CH	4.35, t (6.4)	64.2, CH
4		77.3, C		83.5, C		77.0, C
5		69.7, C		69.5, C		69.8, C
6		65.0, C		74.4, C		66.3, C
7		78.6, C		76.7, C		77.8, C
8	3.09, dd (5.0, 10.2)	18.3, CH2	5.86, d (15.9)	110.5, CH	2.51, dd (17.3, 6.1) 2.57, dd (17.3, 6.3)	24.8, CH2
9	5.50, overlapped	135.3, CH	6.30, dd (5.8, 15.9)	146.3, CH	3.70, td (6.3, 4.3)	71.0, CH
10	5.48, overlapped	124.4, CH	4.00, td (5.8, 1.4)	75.2, CH	3.24, td (6.1, 4.3)	82.1, CH
11	4.39 q (6.7)	67.8, CH	3.47, m	74.4, CH	1.55, m	29.9, CH2
12	1.60, m; 1.44, m	37.4, CH2	1.46, m	33.2, CH2	1.28–1.29, m	25.3, CH2
13	1.27–1.28, m	25.3, CH2	1.27–1.28, m	25.7, CH2	1.28–1.29, m	29.4, CH2
14	1.27–1.28, m	29.3, CH2	1.27–1.28, m	29.4, CH2	1.28–1.29, m	29.9, CH2
15	1.27–1.28, m	31.9, CH2	1.27–1.28, m	31.9, CH2	1.28–1.29, m	31.9, CH2
16	1.27–1.28, m	22.7, CH2	1.27–1.28, m	22.7, CH2	1.28–1.29, m	22.8, CH2
17	0.87 t (6.8)	14.2, CH3	0.87 t (6.9)	14.2, CH3	0.88 t (6.8)	14.2, CH3
−OCH3					3.42, s	58.5, CH3

) showed the presence of two olefinic protons at δ_H 5.50 (1H, m, overlapped, H-9) and 5.48 (1H, m, overlapped, H-10); two oxymethine protons, which appeared at δ_H 4.34 (1H, t, J = 6.4 Hz, H-3) and 4.39 (1H, q, J = 6.7 Hz, H-11); two triplet methyl groups at δ_H 0.99 (3H, t, J = 7.4 Hz, H-1) and 0.87 (3H, t, J = 6.8 Hz, H-17); and fourteen aliphatic methylene protons at δ_H 1.72 (2H, m, H-2), 3.09 (2H, dd, J = 5.0, 10.2 Hz, H-8), 1.60 (1H, m, H-12a), 1.44 (1H, m, H-12b), and 1.27−1.28 (8H, m, overlapped, H-13−16). A detailed analysis of the ¹³C-NMR and DEPT spectra (Table 1) of compound 1 showed signals for two methyl groups at δ_C 9.5 (C-1) and 14.2 (C-17); two O-bearing methine carbons at δ_C 64.1 (C-3) and 67.8 (C-11); two olefinic carbon resonances at δ_C 135.3 (C-9) and 124.4 (C-10); and seven methylene carbons with chemical shifts ranging from 18.3 to 37.4 ppm. The abovementioned signals accounted for one degree of unsaturation, and thus, the remaining required the existence of two additional triple bonds in the molecule, which consisted of four quaternary carbons δ_C 77.3 (C-4), 69.7 (C-5), 65.0 (C-6), and 78.6 (C-7) corresponding to the observed substitutions in the UV and IR spectra. By comparison, these data were relatively close to those of panaxjapyne A [6], except for the additional oxymethine at δ_C 67.8 (C-11), which was clearly proven by the ¹H-¹H COSY correlations (Figure 2) of H-11 with H-10 and H-12 and HMBC correlations (Figure 2) from H-11 to C-9, C-10, C-12, and C-13. Accordingly, compound 1 was elucidated as heptadeca-9-ene-4,6-diyne-3,11-diol.

Compound 2 was separated as a yellow oil with a negative optical rotation ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −80 (c 0.1, CHCl₃), and its molecular formula was determined to be C₁₇H₂₆O₃ by the HRESIMS peak at m/z 301.1776 [M + Na]⁺ (calcd C₁₇H₂₆NaO₃ for 301.1780). The ¹H-NMR (Table 1) spectrum showed a pair of trans olefinic protons at δ_H 6.30 (1H, dd, J = 5.8, 15.9 Hz, H-9) and 5.86 (1H, d, J = 15.9 Hz, H-8); three oxymethine protons at δ_H 4.41 (1H, t, J = 6.5 Hz, H-3), 4.00 (1H, td, J = 5.8, 1.4 Hz, H-10), and 3.47 (1H, m, H-11); and two methyl groups at δ_H 1.01 (3H, t, J = 7.4 Hz, H-1) and 0.87 (3H, t, J = 6.9 Hz, H-17). A detailed analysis of the NMR spectroscopic spectra of compound 2 revealed that it is similar to panaxjapyne B [6] except for the additional oxymethine group in compound 2. The ¹H-¹H COSY correlations (Figure 2) of H-8/H-9/H-10/H-11/H-12 and the HMBC correlations (Figure 2) from H-10 to C-11 and C-12 and from H-11 to C-9 and C-13 evidenced the location of oxymethine at C-11, which accounted for the molecular weight difference of 16 amu observed between the two compounds. Thus, the structure of compound 2 was elucidated as (E)-heptadeca-8-ene-4,6-diyne-3,10,11-triol. The relative configuration of C-10 and C-11 was determined by the ³J_{H, H} value and the ROESY interactions. The small ³J_H-10/H-11 value (1.4 Hz) was indicative of a gauche relationship of H-10 and H-11 [34,35]. The ROESY correlations (Figure 3) of H-9/H-12/H-10/H-11 suggested that the relative configurations of C-10 and C-11 were R* and S*, respectively.

Compound 3 was separated as a yellow oil with a positive optical rotation ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ +80 (c 0.1, CHCl₃). The molecular formula of compound 3 was established as C₁₈H₃₀O₃ from the HRESIMS peak at m/z 317.2089 [M + Na]⁺ (calcd. C₁₈H₃₀NaO₃ for 317.2093). The ¹H and ¹³C-NMR spectroscopic data (Table 1) of compound 3 were comparable to those of oploxyne B [5], suggesting that an oxidized methine of oploxyne B was replaced by the methylene (δ_H 2.51, 2.57/δ_C 24.8) of compound 3. Furthermore, the 16 amu of reduced molecular weight compared with oploxyne B indicated that the molecule formulas of the two compounds only differ by one oxygenium. The ¹H-¹H COSY correlations (Figure 2) of H-8/H-9/H-10/H-11 and the HMBC correlations (Figure 2) from H-8 to C-6, C-7, C-9, and C-10 implied that C-8 is the methylene carbon for compound 3. Finally, the structure of compound 3 was elucidated as 10-methoxyheptadeca-4,6-diyne-3,9-diol.

Compound 4 was obtained as a yellow oil with a negative optical rotation ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −80 (c 0.1, CHCl₃). The molecular formula was determined as C₁₄H₂₂O₃ by the HRESIMS peak at m/z 261.1476 [M + Na]⁺ (calcd for C₁₄H₂₂NaO₃, 261.1467). The ¹H and ¹³C-NMR data (

Table 2. ¹H (500 MHz) and ¹³C-NMR (125 MHz) data for compounds 4 and 5 in CDCl₃ (δ in ppm, J in Hz).

Position	4		5
	δH	δC	δH	δC	Position	δH	δC
1	2.00, s	65.6, CH	2.00, s	66.0, CH	6′		170.1, C
2		68.2, C		68.0, C	7′		170.0, C
3		67.0, C		67.6, C	8′		170.3, C
4		74.5, C		71.6, C	9′		170.6, C
5	2.58, dd (17.4, 6.8) 2.64, dd (17.4, 6.5)	24.6, CH2	2.62, d (5.6)	21.8, CH2	6″	2.11 b, s	20.8 a, CH3
6	4.13, br t (6.5)	69.6, CH	5.11, q (5.6)	69.1, CH	7″	2.12 b, s	21.0 a, CH3
7	3.49, m	73.1, CH	5.39, dd (4.7, 6.2)	70.6, CH	8″	2.10 b, s	20.9 a, CH3
8	3.82, m	75.1, CH	5.18, dd (4.7, 6.2)	71.2, CH	9″	2.10 b, s	20.8 a, CH3
9	1.55, m	33.6, CH2	5.07, q (6.4)	71.5, CH
10	1.33, m1.55, m	26.0, CH2	1.55, m	30.6, CH2
11	1.30, br s	29.4, CH2	1.28, m	24.6, CH2
12	1.30, br s	31.9, CH2	1.28, m	31.5, CH2
13	1.30, br s	22.7, CH2	1.27, m	22.5, CH2
14	0.89, t (6.7)	14.2, CH3	0.86, t (6.9)	14.1, CH3

^a,b exchangeable.

) showed three O-bearing methine groups at δ_H 4.13 (1H, br t, J = 6.5 Hz, H-6)/δ_C (69.6), 3.82 (1H, m, H-8)/δ_C (75.1), and 3.49 (1H, m, H-7)/δ_C (73.1); one triplet methyl group at δ_H 0.89 (3H, t, J = 6.7, H-14)/δ_C (14.2); one acetylene CH group at δ_H 2.00 (1H, s, H-1)/δ_C (65.6); three quaternary carbon groups at δ_C 68.2 (C-2), 67.0 (C-3), and 74.5 (C-4); and six methylenes at δ_H (1.30–2.64)/δ_C (22.7–33.6). The ¹H and ¹³C-NMR spectroscopic data of compound 4 approached those of (6S,7S)-6,7-dihydroxytetradeca-1,3-diyne [36] except for an additional O-bearing methine in compound 4. The structure was further elucidated by the HMBC and ¹H-¹H COSY data (Figure 2). The ¹H-¹H COSY correlations between H-5/H-6/H-7/H-8/H-9, together with the HMBC correlations from H-8 to C-6, C-9 (δ_C 33.6), and C-10 (δ_C 26.0) and from H-6 to C-4 and C-5 (δ_C 24.6) suggested that compound 4 has three successive O-bearing CH groups located at C-6, C-7, and C-8. Thus, the structure of compound 4 was elucidated as tetradeca-1,3-diyne-6,7,8-triol.

Compound 5 was obtained as a yellow oil with a negative optical rotation ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −16 (c 0.1, CHCl₃). The molecular formula was determined as C₂₂H₃₀O₈ by the HRESIMS peak at m/z 445.1859 [M + Na]⁺ (calcd for C₂₂H₃₀NaO₈, 445.1833). The ¹H-NMR spectrum (Table 2) showed four O-bearing methine groups at δ_H 5.11 (1H, q, J = 5.6 Hz, H-6), 5.39 (1H, dd, J = 4.7, 6.2 Hz, H-7), 5.18 (1H, dd, J = 4.7, 6.2 Hz, H-8), and 5.07 (1H, q, J = 6.4 Hz, H-9); one triplet methyl group at δ_H 0.86 (3H, t, J = 6.9 Hz, H-14); one acetylene CH group at δ_H 2.00 (1H, s, H-1); 10 methylene protons at δ_H 1.67–2.62; and 12 acetoxy methyl protons at δ_H 2.10–2.12. The ¹³C-NMR and DEPT spectra indicated the presence of 22 carbons—one terminal methyl carbon at δ_C 14.1 (C-14); four O-bearing methine carbons at δ_C 69.1 (C-6), 70.6 (C-7), 71.2 (C-8), and 71.5 (C-9); three quaternary acetylenic carbons at δ_C 68.0 (C-2), 67.6 (C-3), and 71.6 (C-4); one tertiary acetylenic carbon at δ_C 66.0 (C-1); and five methylene carbons at 21.8–31.5 ppm. All the assignments were supported by the HSQC experiments. A detailed comparison of the spectroscopic data of compounds 4 and 5 showed that in compound 5, there were four more acetyl groups [δ_C (170.0–170.6)/δ_H (2.10–2.12)] and one more O-bearing methine than in compound 4. The ¹H-¹H COSY correlations (Figure 2) of H-5/H-6/H-7/H-8 and H-9/H-10/H-11, as well as the HMBC correlations (Figure 2) from H-6/H-6″ to C-6′ (170.1), from H-7/H-7″ to C-7′ (170.0), from H-8/H-8″ to C-8′ (170.3), and from H-9/H-9″ to C-9 (170.6), indicated that compound 5 has four acetyls linked to C-6, C-7, C-8, and C-9, respectively. The other key correlations of HMBC and ¹H-¹H COSY are shown in Figure 2. Thus, the planar structure of compound 5 was elucidated as 6,7,8,9-tetraacetoxytetradeca-1,3-diyne.

Due to the high flexibly of the carbon chains, crystals were not obtained from compounds 1–5. Furthermore, the triple bonds contained compounds that were unstable and highly reactive [1], which resulted in all the isolates being chemically changed via oxidation and degradation. Therefore, the configurations of compounds 1–5 were not elucidated because of the materials’ instability and were reported as shown in Figure 1.

Compounds 1–7 were assessed for cytotoxic activity in the BEL-7402, K562, and SGC-7901 cancer cell lines, respectively. The results show that compounds 1 and 6 exhibited cytotoxicity in the above three human cancer lines, ranging from 14.3 to 45.4 μM (

Table 3. Cytotoxicity of compounds 1–7 in the human cancer cell lines.

Compound	Cell Line, IC50 (μM)
	BEL-7402	SGC-7901	K562
1	24.9 ± 0.3	45.4 ± 0.6	16.8 ± 0.1
4	>50	26.2 ± 0.4	21.9 ± 0.3
6	14.3 ± 0.4	33.4 ± 0.6	16.6 ± 0.4
2, 3, 5, 7	>50	>50	>50
Paclitaxel a	4.3 ± 0.1	4.3 ± 0.2	8.6 ± 0.1

^a Positive control.

). Contrastingly, compound 4 displayed weak cytotoxicity in SGC-7901 and K562, with IC₅₀ values of 26.2 ± 0.4 and 21.9 ± 0.3 μM, respectively. By comparison, the cytotoxicity in BEL-7401 and SGC-7901 was slightly enhanced when the double bond between C-9 and C-10 in compound 1 was oxidized to the epoxy group in compound 6, which could be due to the highly genotoxic effect of the epoxy group [37,38].

3. Materials and Methods

3.1. General Experimental Procedures

The ¹H, ¹³C, and 2D NMR spectra were recorded on a Bruker AV III spectrometer (Bruker, Bremen, Germany) at either 500 MHz (¹H) or 125 MHz (¹³C) using TMS as an internal standard. The HRMS were measured with an API QSTAR Pulsar mass spectrometer (Bruker). The UV spectra were performed on a Shimadzu UV-2550 spectrometer (Beckman, Brea, CA, USA). The IR absorptions were obtained on a Nicolet 380 FT-IR instrument (Thermo, Pittsburgh, PA, USA) using KBr pellets. The optical rotation was measured on a Rudolph Autopol III polarimeter (Rudolph, Hackettstown, NJ, USA). Silica gel (60–80, 200–300 mesh, Qingdao Marine Chemical Co. Ltd., Qingdao, China), ODS gel (20–45 μm, Fuji Silysia Chemical Co. Ltd., Durham, NC, USA), and Sephadex LH-20 (Merck, Darmstadt, Germany) were used for column chromatography. The TLC was conducted on pre-coated silica gel G plates (Qingdao Marine Chemical Co. Ltd.), and spots were detected by spraying with 10% H₂SO₄ in EtOH followed by heating.

3.2. Plant Material

The plant material was collected at the Chinese Academy of Tropical Agricultural Sciences, Haikou, China, in April 2014 and was identified as Swietenia macrophylla by Dr. Jun Wang (Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences). The voucher specimen (No. DYTHXM201404) was deposited at the Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Science.

3.3. Extraction and Isolation

The roots of S. macrophylla (57.0 kg, dry weight) were first crushed and extracted with 95% EtOH (3 × 150.0 L) at room temperature and evaporated to yield EtOH extract (8.0 kg), which was then partitioned with H₂O (40.0 L) and extracted with petroleum ether (PE) (40.0 L × 3) and EtOAc (40.0 L × 3), respectively. The EtOAc extract (3931.0 g) was subjected to silica gel (20 × 50 cm, 12.0 kg) vacuum liquid chromatography, and eluted with CHCl₃/MeOH (v/v, 10:1; 40.0 L) to obtain fraction 1 (Fr.1). Silica gel (10 × 50 cm, 2.0 kg) vacuum liquid chromatography of Fr.1 (573.0 g) was eluted with PE/EtOAc (v/v, 1:0, 100:1, 50:1, 25:1, 10:1, 5:1, each 5.0 L, gradient) and CHCl₃/MeOH (v/v, 1:0, 100:1, 50:1, 25:1, 10:1, 5:1, 0:1, each 5.0 L, gradient), respectively, which resulted in 17 sub-fractions (Fr.1.1−Fr.1.17). Fr.1.8 (19.5 g) was applied to an ODS gel (4.5 × 40 cm) and eluted with MeOH/H₂O (v/v, 3:7, 2:3, 1:1, 3:2, 7:3, 4:1, 9:1, 1:0, each 5.0 L) to yield Fr.1.8.1−10. Fr.1.8.5 (2.9 g) was separated on a Sephadex LH-20 (3 × 100 cm) with CHCl₃/MeOH as the eluent (v/v, 1:1; 1.5 L) to give Fr.1.8.5.1−5. Fr.1.8.5.1 (1.6 g) was separated on a silica gel column (3 × 45 cm, 160 g) and eluted with PE/EtOAc (v/v, 12:1) to yield compound 1 (30.0 mg) before it was eluted with a gradient of PE/acetone (v/v, 20:1, 15:1, 5:1, 1:1, 0.6 L of each) to give six fractions (Fr.1.8.5.1.1−6). Compound 7 (2.2 mg) from Fr.1.8.5.1.1 (32.0 mg) was obtained from a silica gel (1 × 12 cm, 6 g) eluting with PE/EtOAc (v/v, 8:1). Compound 6 (30.0 mg) from Fr.1.8.5.1.2 (380.0 mg) was obtained from a silica gel (2 × 30 cm, 60.0 g) eluting with PE/CHCl₃ (v/v, 1:1). Fr. 1.8.5.1.3 (150.0 mg) was subjected to a silica gel (1 × 25 cm, 15.0 g) with PE/EtOAc (v/v, 5:1) to obtain compound 3 (8.0 mg). Fr. 1.8.5.1.4 (180.6 mg) was applied to a silica gel (1 × 30 cm, 18.0 g) eluting with PE/EtOAc (v/v, 4:1) to yield compound 2 (14.8 mg). Fr. 1.8.5.1.5 (121.4 mg) was purified by a silica gel (1 × 30 cm, 20.0 g) eluting with CHCl₃/acetone (v/v, 5:1) to obtain compounds 5 (1.2 mg) and 4 (3.8 mg).

Heptadeca-9-ene-4,6-diyne-3,11-diol (1): yellow oil; UV (CH₃OH) λ_max (log ε) 280 (2.90), 270 (2.98) nm; IR (KBr) ν_max 3435, 2924, 2234, 1639, 1384, 1020 cm⁻¹; ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ +10 (c 0.1, CH₃OH); ¹H and ¹³C-NMR data: Table 1; HRESIMS m/z 285.1830 [M + Na]⁺ (calcd. C₁₇H₂₆NaO₂ for 285.1831).

(E)-Heptadeca-8-ene-4,6-diyne-3,10,11-triol (2): yellow oil; UV (CHCl₃) λ_max (log ε) 286 (3.60), 270 (3.68) nm; IR (KBr) ν_max 3433, 2927, 2250, 1645, 1382, 1026 cm⁻¹; ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −80 (c 0.1, CHCl₃); ¹H and ¹³C-NMR data: Table 1; HRESIMS m/z 301.1776 [M + Na]⁺ (calcd. C₁₇H₂₆NaO₃ for 301.1780).

10-Methoxyheptadeca-4,6-diyne-3,9-diol (3): yellow oil; UV (CHCl₃) λ_max (log ε) 286 (2.77), 270 (2.78) nm; IR (KBr) ν_max 3432, 2929, 2245, 1388, 1028 cm⁻¹; ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ +80 (c 0.1, CHCl₃); ¹H and ¹³C-NMR data: Table 1; HRESIMS m/z 317.2089 [M + Na]⁺ (calcd. C₁₈H₃₀NaO₃ for 317.2093).

Tetradeca-1,3-diyne-6,7,8-triol (4): yellow oil; UV (CHCl₃) λ_max (log ε) 298 (1.32), 240 (2.60) nm; IR (KBr) ν_max 3434, 2926, 2241, 1384, 1018 cm⁻¹; ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −80 (c 0.1, CHCl₃); ¹H and ¹³C-NMR data: Table 2; HREIMS m/z 261.1476 [M + Na]⁺ (calcd for C₁₄H₂₂NaO₃, 261.1467).

6,7,8,9-Tetraacetoxytetradeca-1,3-diyne (5): yellow oil; UV (CHCl₃) λ_max (log ε) 212 (4.44), 271 (3.55), 308 (3.12) nm; IR (KBr) ν_max 2924, 2233, 1741, 1634, 1460, 1090 cm⁻¹; ${\left[\alpha \right]}_{\mathrm{D}}^{25}$ −16 (c 0.1, CHCl₃); ¹H and ¹³C-NMR data: Table 2; HREIMS m/z 445.1859 [M + Na]⁺ (calcd for C₂₂H₃₀NaO₈, 445.1833).

3.4. Bioassay of Cytotoxic Activity

MTT assay, originally described by Mosmann [39], was used to quantitate the cytotoxicity of compounds 1–7. The human hepatocellular carcinoma cell line BEL-7402, human myeloid leukemia cell line K562, and human gastric carcinoma cell line SGC-7901, which were obtained from the cell bank of type culture collection of the Chinese Academy of Sciences, Shanghai Institute of Cell Biology, were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum at the conditions of 37 °C, 5% CO₂, and 90% humidity. Paclitaxel was used as the positive control and DMSO was used as the negative control. Different concentrations of the test sample (each had triplicate wells) were designed as 0.1, 0.4, 1.6, 6.3, 25, and 100 μM. The logarithmic phase cells (90 μL) were selected to seed onto the 96-well plates at a concentration of 5 × 10⁴ cell/mL. Then, 15 μL of MTT dissolved in PBS at 5 mg/mL was added to each well, and the system was incubated at 37 °C for 4 h. After that, the supernatant was discarded, and 100 μL of DMSO was added into each well. Finally, the OD value was measured by a MK3 Microtiter plate reader at a wavelength of 490 nm.

4. Conclusions

Seven polyacetylenes were isolated from the roots of S. macrophylla. Their structures were determined by spectroscopic analysis and comparing data in the literature. Furthermore, compounds 1 and 6 displayed weak cytotoxicity in the BEL-7402, SGC-7901, and K562 cell lines, and compound 4 showed a weak cytotoxic effect in the SGC-7901 and K562 cell lines.